The long-term goal of this project is to understand the molecular basis for defining ion selectivity and conductance of paracellular transport and to manipulate them for therapeutic purposes. Transport across epithelia of the GI tract and liver occurs through both a transcellular and paracellular pathway. The intercellular tight junction creates the major barrier in the paracellular pathway. Its properties are highly variable among epithelia in terms of electrical resistance and ion selectivity. The molecular basis for defining these differences remains unknown. Barrier disruption alters absorption/secretion and invasion by pathogens and antigens, thus contributing to a wide range of diseases. The ability to manipulate paracellular properties has significant therapeutic implications. A large family of transmembrane proteins called the claudins forms cell-to-cell contacts at the tight junction and is implicated in creating the variable properties. In preliminary studies, several claudins have been overexpressed in a cultured epithelial cell model and each found to confer a different effect on paracellular conductance and the permeabilities of Na+ and Cl- ions. The present study will test the hypothesis that these channel-like properties are based on the extracellular amino acid sequences of the claudins. Chimeric and site-directed mutant proteins will be expressed in a cultured epithelial cell line and their electrophysiologic properties tested.